title: Trans-scleral imaging of the human trabecular meshwork by two-photon microscopy.
abstract: OBJECTIVE
  To image the native (unfixed) human trabecular meshwork (TM) through the overlying sclera using a non-invasive, non-destructive technique.
  
  
  METHODS
  Two-photon microscopic (2PM) methods, including two-photon autofluorescence (2PAF) and second harmonic generation (SHG), were used to image through the sclera of a human cadaver eye into the TM region. Multiple images were analyzed along the tissue axis (z-axis) to generate a three-dimensional (3D) model of the region. The tissue was subsequently fixed, paraffin embedded, and histological sections were photographed for comparison to the 2PM images.
  
  
  RESULTS
  3D analysis of multiple 2PM SHG images revealed an open region deep within the TM consistent with the location of Schlemm's canal (SC). Images of the scleral spur and surrounding tissues were also obtained. The SC, TM, scleral spur, and surrounding tissue images obtained with 2PM matched with histologically stained sections of the same tissue.
  
  
  CONCLUSIONS
  2PM imaging of the outflow system of the human eye documented collagenous structures solely from inherent optical properties. 2PM successfully imaged through the sclera into the SC/TM without the need for fixation, embedding, or histological processing. This work reveals that 2PM imaging has potential as a new metric for evaluating the aqueous outflow region of the human eye and is worthy of further exploration.
authors.affiliation: Department of Ophthalmology, University of Colorado Denver, Aurora, CO 80045, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Ammar
authors.forename: Tim C
authors.initials: TC
authors.lastname: Lei
authors.forename: Omid
authors.initials: O
authors.lastname: Masihzadeh
authors.forename: Emily A
authors.initials: EA
authors.lastname: Gibson
authors.forename: Malik Y
authors.initials: MY
authors.lastname: Kahook
mesh_terms: D000368:Aged; D006801:Humans; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008297:Male; D036641:Microscopy, Fluorescence, Multiphoton; D012590:Sclera / Q000166:cytology*; D014129:Trabecular Meshwork / Q000166:cytology*